Ghrelin is a bioactive peptide that induces food intake, body weight gain, and adiposity in rodents (Tschop M. et al., Nature 407:908-13 (2000); Wren A. M. et al., Diabetes 50:2540-47 (2001)). Acute administration of ghrelin induces food intake in healthy men and women (Wren A. M. et al., J. Endocrinol. Metab. 86:5992-95 (2001); Druce M. R. et al., Int. J. Obes. Relat. Metab. Disord. 29:1130-36 (2005)).
The GHRL (ghrelin) gene encodes a variety of products resulting from alternatively spliced transcripts, various types of cleavage of the prepropeptide, and several post-translational modifications (Kojima M. & Kangawa M., Physiol. Rev. 85:495-522 (2005); Zhang J. V. et al., Science 310:996-99 (2005)). In addition, different degradation products are produced in various tissues (De Vriese C. at al., Endocrinology 145:4997-5005 (2004)). Some of these GHRL products are described herein.
Ghrelin is a 28 amino acid peptide bearing an n-octanoyl side chain on the third serine, resulting from the cleavage of signal and propeptide from the 117 amino acid preproghrelin and an acylation event. The acylated N-terminus of ghrelin is essential for the endocrine functions (Kojima M. at al., Nature 402:656-60 (1999); Bednarek M A et al., J. Med. Chem. 43:4370-76 (2000)). Des-acyl ghrelin, which lacks the endocrine functions, was shown to have an antagonistic effect to that of ghrelin on glucose output in vitro (Gauna C. at al., J. Clin. Endocrinol. Metab. 89:5035-42 (2004)). An alternatively-spliced ghrelin mRNA encodes a 116 amino acid prepropeptide that is further processed to a Des-Gln14-ghrelin and a 27 amino acid processed peptide (Hosoda H. at al., J Biol. Chem. 275:21995-22000 (2000)). Another peptide, Obestatin, is cleaved from the preproghrelin and has no sequence overlap with processed ghrelin peptide. It was shown to have some antagonistic effect to acylated ghrelin, inhibiting food intake and body weight gain (Zhang J. V. at al., Science 310:996-99 (2005)). Yet another peptide, the 66 amino acid C-terminus of the preproghrelin, may also be functional (Pemberton C. at al., Biochem. Biophys. Res. Comm. 310:567-73 (2003)). A variety of isoforms, including isoforms encoded by different splice variants, are known for other proteins, e.g. for vascular endothelial growth factor (VEGF) where different isoforms share roles as angiogenesis, while differing in some other characteristics as binding affinity (Neufeld G. et al., FASEB J. 13:9-22 1999). Thus, the variety of products of the GHRL gene may reflect a similarly complex control of the endocrine and paracrine action of the ghrelin isoforms.
Ghrelin was previously shown to specifically bind a species of high density lipoprotein associated with the plasma esterase, paraoxonase, and clusterin. An endogenous species of ghrelin was found to co-purify with high density lipoprotein during density gradient centrifugation and subsequent gel filtration. This interaction links the orexigenic peptide hormone ghrelin to lipid transport and metabolism (Beaumont J. et al., J. Biol. Chem. 11:8877-80 (2003)).
Research from experimental animals, laboratory investigations, epidemiology and genetic forms of hypercholesterolemia indicate that elevated cholesterol is a major cause of CHD (Coronary Heart Disease). The Framingham Heart Study also established that high blood cholesterol is a risk factor for CHD. Results of the Framingham study showed that the higher the cholesterol level, the greater the risk of suffering CHD. On the other end of the spectrum, CHD is uncommon at total cholesterol levels below 150 milligrams per deciliter (mg/dL). A direct link between high blood cholesterol and CHD has been confirmed by the Lipid Research Clinics-Coronary Primary Prevention Trial (1984), which showed that lowering total and LDL cholesterol levels significantly reduces CHD. A series of more recent trials of cholesterol lowering using statin drugs has demonstrated conclusively that lowering total cholesterol and LDL-cholesterol reduces the chance of suffering a heart attack, needing bypass surgery or angioplasty, and dying of CHD-related causes.
Ghrelin is synthesized in the stomach (as well as in the intestine, pituitary gland, and possibly in the hypothalamus) and activates the growth hormone secretagogue receptor. Ghrelin secretion increases with decreased food intake in animals and humans and stimulates food intake. Thus, the “drive to eat” after dieting may be partially because of ghrelin secretion. Reducing ghrelin activity may reduce the “drive to eat,” and, in fact, it has been suggested that it is the reduction in ghrelin that partially accounts for the effectiveness of gastric bypass surgery (Cummings D E et al., N. Eng. J. Med. 346(21):1623-30 (2002)). Therefore, ghrelin antagonism may potentially decrease or at least blunt the increased appetite that may occur with decreased feeding and, thus, be a potential adjunctive treatment for obesity.
Obesity is the most common metabolic disease in developed nations. Despite public health education and initiatives, its prevalence continues to increase, with 30% of adults in the United States being obese and 60% of adults being overweight or obese. The World Health Organization has estimated that worldwide, over one billion adults are overweight, with at least 300 million of them being obese. The increasing prevalence of obesity among children and adolescents is also of great concern and suggests a likelihood of worsening obesity trends in future adults. Obesity leads to, or significantly increases the risk of, comorbidities involving various body systems including complications in the following systems: 1) cardiovascular [hypertension, congestive cardiomyopathy, varicosities, pulmonary embolism, coronary heart disease (CHD)1], 2) neurological (stroke, idiopathic intracranial hypertension, meralgia parethetica), 3) respiratory (dyspnea, obstructive sleep apnea, hypoventilation syndrome, Pickwickian syndrome, asthma), 4) musculoskeletal (immobility, degenerative osteoarthritis, low back pain), 5) skin (striae distensae or “stretch marks,” venous stasis of the lower extremities, lymphedema, cellulitis, intertrigo, carbuncles, acanthosis nigricans, skin tags), 6) gastrointestinal (GI; gastro-esophageal reflux disorder, nonalcoholic fatty liver/steatohepatitis, cholelithiasis, hernias, colon cancer), 7) genitourinary (stress incontinence, obesity-related glomerulopathy, breast and uterine cancer), 8) psychological (depression and low self-esteem, impaired quality of life), and 9) endocrine (metabolic syndrome, type 2 diabetes, dyslipidemia, hyperandrogenemia in women, polycystic ovarian syndrome, dysmenorrhea, infertility, pregnancy complications, male hypogonadism). Therefore, it has been a therapeutic and scientific goal to develop strategies to reduce the worldwide obesity epidemic to develop safe and effective antiobesity drugs, analogous to the development of treatments against hypertension, dyslipidemia, and diabetes.